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spinqualitylabelsall models Enoyl-ACP Reductase InhA Homotetramer Show:Asymmetric Unit Biological Assembly Drag the structure with the mouse to rotate   Export Animated Image

Contents 1 Introduction 1.1 FAS-II System 1.2 Mechanism of Action 2 General Structural Information 2.1 Ligands 2.2 Fatty Acyl Binding Crevice 2.2.1 Importance of Tyr-158 2.3 Catalytic Triad 2.3.1 Hydrogen Bonding Interactions 3 Clinical Applications

Introduction

FAS-II System

Mechanism of Action

General Structural Information

Crystal structures of InhA reveal a (each subunit featured with a different color) in aqueous solution with separate ligand binding sites in each subunit. Each subunit is composed of 289 residues and features a typical Rossmann fold containing a single NADH binding site. The of InhA is made up of several alpha helices (pink), beta sheets (gold), and beta turns (white). This enzyme also features a fatty acyl binding crevice that accommodates the long-chain fatty acyl substrate needed to synthesize mycolic acid precursors. The of the InhA form one side of the fatty acyl binding crevice, referred to as the (residues 196-219). One side of this crevice is open and exposed to solvent, which allows the substrates to access the binding pocket of this enzyme. The size of the substrate binding loop is a primary determinant of the ability of InhA to select for fatty acyl chains longer than 16 carbons to successfully produce mycolic acid precursors.

Talk generally about catalytic triad. More specific details in sections below.

Ligands

The two ligands that play a role in the function of InhA are NADH and the long-chain fatty acyl substrate. NADH, or nicotinamide adenine dinucleotide, is a cofactor found in all living cells. It is especially important in the function of InhA because it helps position the fatty acyl substrate within the fatty acyl binding crevice, thereby improving the overall activity of InhA. Once properly positioned, the fatty acyl substrates are reduced at the trans double bond between C2 and C3, forming mycolic acid precursors that eventually compose the cell wall of mycobacteria.

Fatty Acyl Binding Crevice

Within the fatty acyl binding crevice, the NADH substrate sits on the top shelf of the Rossmann fold, and the fatty acyl substrate sits on top of the NADH substrate. Due to its position within the crevice, the trans double bond of the fatty acyl substrate is found near the closed end of the crevice and is located directly adjacent to the nicotinamide ring of NADH. These two molecules interact with each other via hydrogen bonding. The first hydrogen bond occurs between the phosphate oxygen of NADH and the amide nitrogen of the N-acetylcysteamine portion of the fatty acyl substrate. The second hydrogen bond occurs between the 2'-hydroxyl of the nicotinamide ribose of NADH and the thioester carbonyl oxygen of the fatty acyl substrate. Additionally, both of these substrates are held in place within the crevice through interactions with the side chains of surrounding (purple) residues. The majority of these residues anchoring the substrates are found within the substrate binding loop itself, including Ala-198, Met-199, Ala-201, Ile-202, Leu-207, Ile-215, and Leu-218. Additional hydrophobic amino acids that are not a part of the substrate binding loop also play a role in positioning and stabilizing the fatty acyl chain in the crevice. Studies have found that the fatty acyl substrate adopts a u-shaped conformation to facilitate binding.

Importance of Tyr-158

One example of a hydrophobic amino acid that is not a part of the substrate binding loop yet interacts with the fatty acyl substrate is Tyr-158. This amino acid is conserved in other enoyl-ACP reductases in both bacteria and plants, so it likely plays an essential role in the function of these specific enzymes. Studies have shown that Tyr-158 forms the only direct hydrogen bond that exists between the InhA protein and the fatty acyl substrate. This hydrogen bond occurs between the hydroxyl oxygen on the side chain of Tyr-158 and the thioester carbonyl oxygen of the fatty acyl substrate. As a result of the highly conserved nature and the specific hydrogen bonding capabilities of this amino acid, it is likely that Tyr-158 plays an important role in fatty acyl substrate binding in enoyl-ACP reductases.

Catalytic Triad

Hydrogen Bonding Interactions

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Clinical Applications

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